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2015 Abstracts

Expression of the C-terminal Domains of the Tight Junction Proteins Claudin-16, -3, and -4 to Identify Interacting Proteins in Epithelial Ovarian Carcinoma

Brandon Davies, Utah Valley University

Physical Sciences

Epithelial ovarian carcinoma (EOC) is the sixth most common cancer in US women. The long-term cure rates are low due to the lack of reliable biomarkers for early disease detection, resulting in advanced stage diagnosis. Approximately 75%-80% of ovarian cancers are diagnosed at stages IIIV with a 10% 5-year survival rate despite aggressive treatments. Claudin proteins are being studied as possible biomarkers as they are aberrantly overexpressed in EOC tumors. The Claudin family of proteins are a main component of tight junctions in the upper region of epithelial cells that act as gateways for the exchange of water and solutes while also helping determine the cell’s polarity and function. Changes in these proteins cause changes in phenotype and function of normal epithelial cells, such as proliferation control, trans-epithelial resistance, polarity, and solute transport. Claudin-16 is often aberrantly expressed in breast and ovarian cancer, while Claudins 3- and 4 are highly overexpressed in EOC. The location of these proteins is also correlated with oncogenic transformations and cell proliferation. Determining the specific characteristics of these Claudin proteins can prove to be of incredible benefit in cancer treatments. As these proteins are targeted during these therapies, these tight junctions may then send normal signals, which in turn can regulate the cell normally. The C-termini of the Claudins, which are cytoplasmically located, contain a known PDZ-binding motif and may interact with other junction proteins or with proteins involved in interesting signaling pathways. To identify these interacting proteins, we will use the Expresso T7 Cloning System (Lucigen Corp., Middleton, WI) to purify the Claudin-16, -3, and -4 C-terminal tails to use in pull-down assays. This process includes using affinity tags to capture the Claudin tails by FPLC, which can then be analyzed by SDS-PAGE and, ultimately, the corresponding genes cloned and sequenced. This study can potentially provide crucial information in relation to how members of the Claudin family interact with other proteins that are commonly found in tissues that are misregulated in cancer. With this data treatments can be improved to increase the responsiveness of ovarian cancer patients.